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Abstract

Current optical super-resolution implementations are capable of resolving features spaced just a few nanometers apart. However, translating this spatial resolution to cellular targets is limited by the large size of traditionally employed primary and secondary antibody reagents. Recent advancements in small and efficient protein binders for super-resolution microscopy such as nanobodies or aptamers provide an exciting avenue for the future, however their widespread availability is still limited. To address this issue, we here report the combination of bacterial-derived binders commonly used in antibody purification with DNA-PAINT microscopy. The small size of these protein binders compared to secondary antibodies make them an attractive labeling alternative for emerging super-resolution techniques. We here present a labeling protocol for DNA conjugation of bacterial-derived protein A and G for DNA-PAINT imaging and assay their performance intracellularly by targeting primary antibodies against Tubulin, TOM20, and EGFR and quantify the increase in obtainable resolution. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.